This project is concerned with the mechanisms of action and regulation of glucose 6-phosphate dehydrogenases (G6PD). Most of the studies concern G6PD from lactating rat mammary gland and from Leuconostoc mesenteroides. The mammary G6PD exists in several forms resolvable on polyacrylamide gel electrophoresis. We will attempt to characterize the structural and kinetic differences between these. For the bacterial enzyme we are continuing studies on the identification of the nature and function of active site amino acids; the determination of binding constants for the coenzymes and their analogs; monitoring conformational changes associated with binding of various ligands; elucidation of the stereospecificity of H-transfer; and elucidation of the nature of the multiple electrophoretic forms of this enzyme. We are also comparing the molecular weights and polyacrylamide gel electrophoresis patterns of G6PD's from several rodent, bovine and human tissues.